When successful, bone marrow transplantation (BMT) is a cure for non-malignant hematological disorders, including: sickle cell disease, aplastic anemia, thalassemia major, Diamond-Blackfan Anemia and Fanconi Anemia. However, most patients who suffer from the above illnesses receive chronic red blood cell (RBC) transfusions, as a life-sustaining measure. BMT is difficult to achieve in chronically transfused patients, who have graft rejection at a much higher frequency than with BMT in other settings. This severely limits the feasibility of utilizing BMT as a cure for the above disorders. A leading hypothesis to explain the increased rate of BMT rejection is that multiple transfusions result in immunization to transplantation antigens. Since BMT in this setting is MHC matched, these antigens would be minor histocompatibility antigens (mHAs). Previously, it has been assumed that contaminating leukocytes in blood transfusions were the main source of immunization. However, the implementation of stringently leukoreduced blood products (fewer than 1x106 total leukocytes per unit of blood) has not decreased the rate of BMT rejection in chronically transfused patients. Based upon these findings, we hypothesized that transfusion of red blood cells (RBCs) is alone sufficient to immunize against mHAs. In support of this concept, we recently reported a novel mechanism by which mHAs on transfused RBCs are crosspresented into the MHC class I pathway of recipient antigen presenting cells (APCs), resulting in activation and expansion of recipient CD8+ T cells specific for the mHAs. Moreover, recent experiments in our lab have shown that transfusion of leukoreduced RBCs leads to sufficient alloimmunization against mHAs to induce BMT rejection. In this context, we propose to study the mechanisms of RBC transfusion induced BMT rejection. Specific Aim 1: Elucidate the immune mechanisms of RBC transfusion-induced BMT rejection. Specific Aim 2: Analysis of CD4+ T and CD8+ T cell immunization by mHAs on transfused RBCs. Specific Aim 3: Analysis of Antigen Presentation Cells in Transfusion-Induced BMT Rejection. Together, the proposed aims will provide a mechanistic elucidation of how RBC transfusion induces subsequent BMT rejection. These studies have the potential to directly benefit patient populations with non- malignant hematological disorders (i.e. sickle cell anemia), as generating strategies to prevent RBC alloimmunization may allow successful implementation of BMT as a cure for these diseases. PUBLIC HEALTH RELEVANCE: When successful, bone marrow transplantation is a cure for a variety of diseases that involve a defect in blood cell production, including: sickle cell anemia, idiopathic aplastic anemia, thalassemia major, Diamond-Blackfan Anemia and Fanconi Anemia. However, several barriers prevent the implementation of bone marrow transplantation as a cure in this population. The proposed studies investigate one of the main barriers to bone marrow transplantation in an effort to understand and overcome the problems of bone marrow transplantation in the above patient population. Thus, the proposed studies are relevant to the treatment/cure of a variety of diseases involving blood cell production.